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Membrane Traffic Flashcards

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1
Q

Functions of the endoplasmic reticulum (RER and SER -3)

A
  1. synthesis of cholesterol, phospholipids, triglycerides
  2. accumulation and transport of ions
  3. metabolism of steroids
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2
Q

Functions of the RER only (3)

A

1-3. Translocation, glycosylation and folding of proteins destined for secretion, or export to the golgi, plasma membrane, endosomes and lysosomes

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3
Q

Functions of the golgi complex (7)

A
  1. Protein and lipid sorting
  2. proteoglycan assembly
  3. Glycolipid synthesis
  4. N-linked oligosaccharide processing
  5. O-glycosylation
  6. Sugar phosphorylation
  7. Proteolytic processing
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4
Q

Endocytosis functions (3)

A
  1. Nutrition via receptor-mediated internalization
  2. Homeostasis via regulation of signal transduction and sampling extracellular fluid
  3. Defense - clearing pathogens and debris
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5
Q

Major site of macromolecule degradation

A

lysosome

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6
Q

Lysosome functions (2)

A
  1. Degradation of macromolecules

2. Scaffolding of signaling components (mTORC1)

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7
Q

Common features of secretory pathway signal sequences (3)

A
  1. 1 or 2 basic residues
  2. Hydrophobic
  3. Cleavage site of signal peptidase
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8
Q

Five steps of protein translocation through ER

A
  1. SRP recognizes signal sequence of growing peptide
  2. SRP binding causes a pause in translation
  3. SRP + protein/ribosomes are targeted to ER, translation continues and translocation begins
  4. SRP/ribosomes are released
  5. SRP is recycles
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9
Q

Signal recognition particles - how do they work?

A

Have a signal-sequence binding pocket that attaches to the signal sequence, a hinge that allows for conformational change and targeting, and a translational pause domain that halts translation whilst protein is being targeted to ER

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10
Q

What three things does SRP do?

A
  1. Bind to the signal sequence
  2. Cause a pause or arrest in translation
  3. Bind to the SRP-receptor in the ER membrane
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11
Q

What is the advantage of the SRP-induced translation pause or block?

A

Prevents folding so protein can be translated through Sec61 channel

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12
Q

How does Sec61 keep its pore closed until its needed for translocation?

A

It is constitutively closed by a plug until a signal sequence displaces it

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13
Q

What additional machinery does post-translation translocation require?

A

Sec complex, ATP hydrolysis to phosphorylate BiP

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14
Q

Give an example of a type I ER protein

A

Examples include: glycophorin, LDL receptor, Influenza HA protein, insulin receptor, and growth hormone receptor.
(N terminus on inside, single transmembrane domain, C terminus on outside)

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15
Q

Give an example of a type II ER protein

A

Examples include: transferrin receptor, golgi sialyltransferase, influenza HN protein
(C terminus on inside, single transmembrane domain, N terminus on outside)

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16
Q

Give an example of a type III ER protein

A 
Cytochrome P450
(Anchored transmembrane domain, C terminus outside)
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17
Q

Give an example of a polytopic ER protein

A

GPCRs, glucose transporters, CFTR, Sec61

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18
Q

T/F. Signal sequence cleavage happens after translation

A

False, it is co-translational

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19
Q

How are signal seqs removed from ER membrane

A

Through intramembrane proteolysis, happens rapidly

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20
Q

How does BiP contribute to protein folding?

A

BiP binds hydrophobic patches of protein and acts as barrier to nonspecific binding, as it folds then it kicks off BiP, preventing further binding

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21
Q

What is the role of PDI in ER lumen protein folding?

A

PDI catalyzes the rearrangement of disulfide bonds until the “correct” bond is formed, after which point the protein can proceed to the golgi

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22
Q

Pathway for insertion of tail anchored proteins

A

GET, GET1-2 are transmembrane proteins that guide insertion of tail anchor into membrane through ATPase activity

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23
Q

How is a protein added to a GPI anchor?

A

Nucleophilic displacement reaction involving the attack of NH2 on protein transmembrane domain

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24
Q

What is an example of a mechanism through which misfolded proteins are retrotranslocated or degraded

A

Misfolded proteins bound by lectins, these target to translocator complex, after the protein comes out of this it is targeted for ubiquitination and send to the proteosome

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25
The topology of a plasma membrane protein is determined during:
Synthesis in the ER membrane
26
T/F. ER lumen is the topological equivalent of inside the cell
False, outside the cell
27
Approaches to determining membrane protein topology (4)
1. Standard sequential treatment of +/- membrane, +/- protease, +/- glycosidase and +/- detergent 2. Accessibility of antibody epitopes (ex cell surface staining) 3. Epitope tagging 4. Addition of N-linked glycosylation sites
28
Through which organelle do vesicles travel between the ER and rest of cell?
Golgi apparatus
29
T/F. Membrane topology is maintained during vesicle transport
True
30
Steps of vesicular transport (3) and the types of proteins involved with each step
1. Budding (coats, cargo receptors) 2. Targeting (Tethers, targeting receptors) 3. Fusion (Fusion proteins)
31
Coat complexes used at different steps (3)
1. Clathrin used in vesicles moving from agolgi to endosomes 2. COPI used to coat golgi cisternae 3. COPII used to coat ER vesicles heading to golgi
32
What 4 things are Rab proteins implicated in -
vesicle budding, cargo recruitment, motor recruitment and productive targeting
33
How do Rab proteins promote vesicle targeting
By tethering vesicle to rab effector tethering protein, which brings vesicle into proximity of SNARE complex to ensure proper targeting
34
ATPase required for SNARE disassembly
NSF
35
T/F. NSF and SNAPs required for all vesicle transport steps
True
36
Draw constitutive transport to the plasma membrane mechanistic steps
1. COPII vesicle starts forming with Sar1-GTP bound which starts membrane deformation 2. Sar1 binds Sec23, then cargo binds Sec24. At this point Sec13/31 forms outer shell 3. COPII vesicle formed, with exit signals on both cargo protein and cargo receptor. Only properly folded and assembled cargo is incorporated into COPII vesicles 4. Uncoating with GTP hydrolysis, Sec23 is GAP for Sar1 5. Fusion with cis Golgi network, Rabs/tethers involved 6. Transport through Golgi complex 7. Exit from trans golgi network and fusion with plasma membrane
37
What types of macromolecules are processed in the golgi complex?
N-linked oligosaccharides
38
Purpose of coating proteins (2)
To help deform membrane and select cargo
39
Two models for cargo movement through golgi complex
Vesicular transport model Cisternal maturation model Be able to describe difference
40
Localization signal for ER resident proteins
KDEL (Lys-Asp-Glu-Leu), binds to receptor that internatlizes COP1
41
Localization signal for ER membrane proteins
KKXX dilysine, which binds directly to COP1
42
How are proteins sent where they are supposed to go within golgi?
Through the action of golgi resident glycosyltransferases, which have transmembrane domains but no consensus sequence - and TGN residents which has a Tyr-based signal in cytoplasmic tail
43
Examples of diesases caused by lysosomal storage disorders
Tay-Sachs, Mucolipidosis II, Gaucher's
44
Two types of secretory pathways
Constitutive, all cells have | Regulated, specialized cells such as neurons
45
Some specialized cells which perform regulated secretion
1. Exocrine pancreas, stimulus = acetylcholine, produce = digestive hormones 2. Endocrine pancreas, stimulus = glucose, product = insulin
46
Stimulus secretion coupling steps (3)
1. Secretogogues (stimulus) binds cell surface receptors 2. Second messenger (usually Ca2+) comes in 3. Pre-formed and pre-loaded secretion granules fuse with plasma membrane Time scale = msec (neuron) to minutes (mast cell)
47
Neurotransmitter release at the synapse steps
1. Delivery of synaptic vesicle components to plasma membrane 2. Endocytosis of synatpic vesicle components to form new guys directl;y 3. Endocytosis of components and then delivery to endosome 4. Budding of synaptic vesicle from endosome 5. Loading of neurotransmitter into synaptic vesicle 6. Secretion of neurotransmitter by exocytosis in response to an action potential
48
Sorting in polarized epithelia two types
1. Direct sorting in the trans golgi network | 2. Indirect sorting via endosomes
49
Purpose of endocytosis (3)
Nutrition, defense, homeostasis
50
Types of endocytosis (5)
Phagocytosis, macro-pinocytosis, clathrin mediated or caveolin mediated endocytosis, or pinocytosis not mediated by either
51
Clathrin mediated endocytosis pathway
LDL receptor example - receptor bound to ligand interacts with clathrin coat proteins, eventually pinching off to form vesicle. Vesicle is uncoated, then fuses with early endosome, which matures into a late endosome and then fuses with a lysosome to become an endolysosome
52
AP adapter complexes
have clathrin binding and receptor binding subunits, recognize signals recognized on receptor tails like Tyr-XX-omega, diluecine
53
Endocytosis signal
Essential for internalization, Tyr based signal
54
Other protein required in clathrin-coated vesicle formation
dynamin
55
What determines whether internalized receptors are recycled or degraded?
Sequences in receptor tails, pH at which ligands dissociate
56
Multi-vesicular body formation
Formed through invagination and pinching off of maturing endosome, fusion with lysosome allows you to degrade vesicular contents very efficiently
57
Complexes required for formation of multi-vesicular bodies
ESCRT complex
58
Unique feature of caveolae coat proteins
have hairpin structures that are associated with many signaling proteins, may be involved in signal transduction
59
Steps of phagocytosis
1. Attachment, direct binding or receptor-antibody binding 2. Engulfment (actin assembly) 3. Phagosome formation 4. Degradation (after lysosome fusion)
60
Kinase important for actin remodeling in phagocytosis
PIP3 kinase
61
Strategies for subversion of phagocytosis (3) and an example of an organism which does each
1. Escape from endosome (Trypanosoma cruzi) 2. Prevent fusion with lysosomes (Plasmodium falciparum) 3. Survive in phagolysosome (Leishmania)
62
What is required for clathrin-mediated endocytosis but not phagocytosis
Clathrin

CSD (13 decks)

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